Obesity is a global pandemic with accelerating trends in morbidity, mortality, and medical expenditures. Treatments have been limited in efficacy, or burdened with safety concerns. In the context of this unmet clinical need for more effective and safe anti-obesity therapies, the recent discovery of a new gut-brain endocrine axis regulating appetite and metabolism offers a unique opportunity to advance treatment and prevention for this disease. The transmembrane receptor guanylyl cyclase C (GUCY2C) is well characterized as a key regulator of intestinal epithelial homeostasis upon activation by its cognate paracrine hormones uroguanylin (small intestine) and guanylin (colorectum). The recent discovery of GUCY2C expression and function in hypothalamus, along with circulating uroguanylin levels that increase following caloric intake, extends GUCY2C signaling to the central nervous system in an endocrine regulatory circuit mediating satiety. Indeed, these observations suggest a physiological model in which ingestion of food stimulates the endocrine secretion of uroguanylin from intestine, which circulates to hypothalamus to induce anorexigenic signaling. Further, suppressed postprandial uroguanylin secretion in obese mice suggests a pathophysiologic, but therapeutically targetable, mechanism driving obesity. The ultimate, long-term goal of this project is to elucidate mechanisms by which GUCY2C signaling in hypothalamus regulates satiety, to inform the utility of GUCY2C ligand therapy in the treatment and prevention of obesity. To achieve this long-term objective, three specific aims are proposed. In Aim 1, we will map GUCY2C expression regulating appetite in hypothalamus. Leveraging unique -galactosidase GUCY2C reporter mice, we will determine the hypothalamic nuclei, cell types, and neuronal subtypes expressing GUCY2C. These results will be confirmed by in situ RNA hybridization, immunoblot, immunostaining and GUCY2C signaling analyses. In Aim 2, we will determine the relative roles of hypothalamic and intestinal GUCY2C in regulating appetite. Using tissue-specific conditional mouse models developed in our laboratory, we will eliminate GUCY2C expression in either intestine or brain and determine the effects of these treatments on GUCY2C ligand-induced satiety, as well as chronic effects on food consumption and body weight. In Aim 3, we will define the mechanisms linking GUCY2C activation to satiety signaling. Preliminary data suggests that GUYC2C ligand-induced satiety is associated with increases in expression of hypothalamic POMC, but not other anorexigenic or orexigenic, transcripts. Here, we will demonstrate the role of POMC in linking GUCY2C hypothalamic signaling with regulation of satiety and appetite. Together, these studies will define a novel endocrine mechanism contributing to the normal physiologic control of food intake. They represent the requisite evidence base to ultimately translate this previously unrecognized gut-brain axis into new therapeutic approaches targeting GUCY2C to prevent and treat obesity and metabolic diseases.